To determine the relationship between cerebral Glc metabolism and glutamatergic neuronal function, we used 13 C NMR spectroscopy to measure, simultaneously, the rates of the tricarboxylic acid cycle and Gln synthesis in the rat cortex in vivo. From these measurements, we calculated the rates of oxidative Glc metabolism and glutamate-neurotransmitter cycling between neurons and astrocytes (a quantitative measure of glutamatergic neuronal activity). By measuring the rates of the tricarboxylic acid cycle and Gln synthesis over a range of synaptic activity, we have determined the stoichiometry between oxidative Glc metabolism and glutamateneurotransmitter cycling in the cortex to be close to 1:1. This finding indicates that the majority of cortical energy production supports functional (synaptic) glutamatergic neuronal activity. Another implication of this result is that brain activation studies, which map cortical oxidative Glc metabolism, provide a quantitative measure of synaptic glutamate release.Glc metabolism is the major pathway of energy production in the mature brain (1). During brain activation, increases in Glc metabolism directly form the basis of brain functional mapping by using both 2-deoxyglucose autoradiography (2, 3) and positron-emission tomography (4) and indirectly influence signal changes observed with functional MRI (5). Despite the extensive use of these methods for mapping brain function, the mechanism linking Glc metabolism and functional neuronal activity and the fraction of cerebral energy production that supports neuronal function are still unknown.Glutamate is the major excitatory neurotransmitter in the brain (6), and a high percentage of cortical neurons are glutamatergic (7). It has been proposed that a neuronalastrocytic neurotransmitter cycle exists in the brain in which glutamate from the neuronal pool is released into the synaptic cleft as a neurotransmitter, taken up by astrocytes, converted to Gln, and returned to the neuron in this synaptically inactive form where it is converted back to glutamate (6). The development of in vivo 13 C NMR spectroscopy has enabled the direct investigation of cerebral glutamate metabolism (8, 9). We recently have shown that the rate of glutamateneurotransmitter cycling between neurons and astrocytes can be calculated by using the flux of the 13 C label from glutamate to Gln in the rat brain in vivo during a [1-13 C]Glc infusion (10). Thus, we can obtain an in vivo measure of glutamatergic neuronal activity. In the same experiment, the flux of the 13 C label from [1-13 C]Glc into glutamate yields a simultaneous in vivo measurement of the cerebral tricarboxylic acid (TCA) cycle rate, from which oxidative Glc consumption can be derived (11,12). Therefore, by using the combined measurement of these two fluxes, we can determine quantitatively the stoichiometry between cerebral Glc metabolism and glutamatergic-synaptic activity in vivo.In the present study, we have used direct 13 C NMR spectroscopy to determine the cerebral (primarily cortical)...

Macromolecule resonances underlying metabolites in 1H NMR spectra were investigated in temporal lobe biopsy tissue from epilepsy patients and from localized 1H spectra of the brains of healthy volunteers. The 1H NMR spectrum of brain tissue was compared with that of cytosol and dialyzed cytosol after removal of low molecular weight molecules (< 3500 daltons) at 8.4 and 2.1 Tesla. The assignment of specific resonances to macromolecules in 2.1 Tesla, short-TE, localized human brain 1H NMR spectra in vivo was made on the basis of a J-editing method using the spectral parameters (delta, J) and connectivities determined from 2D experiments in vitro. Two prominent connectivities associated with macromolecules in vitro (0.93-2.05 delta and 1.6-3.00 delta) were also detected in vivo by the J-editing method. Advantage was taken of the large difference in measured T1 relaxation times between macromolecule and metabolite resonances in the brain spectrum to acquire 'metabolite-nulled' macromolecule spectra. These spectra appear identical to the spectra of macromolecules isolated in vitro.

Localized 'H NMR spectroscopy in conjunction with J editing was used to measure the concentration of -aminobutyric acid (GABA) in the occipital lobe of four control human volunteers and four epileptic volunteers who were receiving the drug vigabatrin. The GABA concentration measured in four nonepileptic subjects was 1.1 ± 0.1 #mol/cm3 of brain, which is in good agreement with previous values measured in surgicaly removed human cortex. A dosedependent elevation of GABA concentration was measured in patients receiving the GABA tra inhibitor vigabatrin, with the maximum measured level of 3.7 pmol/cm3 of brain measured at the highest dose (6 g per day) studied. 'H NMR measurements of GABA in those patients receiving GABA-elevating agents such as vigabatrin wiUl be of importance in establishing the relationship between seizure suppression and the concentration of brain GABA.

Recent 13 C NMR studies in rat models have shown that the glutamate͞glutamine cycle is highly active in the cerebral cortex and is coupled to incremental glucose oxidation in an Ϸ1:1 stoichiometry. To determine whether a high level of glutamatergic activity is present in human cortex, the rates of the tricarboxylic acid cycle, glutamine synthesis, and the glutamate͞glutamine cycle were determined in the human occipital͞parietal lobe at rest. During an infusion of [1-13 C]-glucose, in vivo 13 C NMR spectra were obtained of the time courses of label incorporation into [4-13 C]-glutamate and [4-13 C]-glutamine. Using a metabolic model we have validated in the rat, we calculated a total tricarboxylic acid cycle rate of 0.77 ؎ 0.07 mol͞min͞g (mean ؎ SD, n ‫؍‬ 6), a glucose oxidation rate of 0.39 ؎ 0.04 mol͞min͞g, and a glutamate͞ glutamine cycle rate of 0.32 ؎ 0.05 mol͞min͞g (mean ؎ SD, n ‫؍‬ 6). In agreement with studies in rat cerebral cortex, the glutamate͞glutamine cycle is a major metabolic f lux in the resting human brain with a rate Ϸ80% of glucose oxidation.The regulation of the release and re-uptake of the excitatory neurotransmitter glutamate is critical for mammalian brain function (1, 2). Glutamate released from the neuron may be cleared from the synaptic cleft through uptake by neuronal or glial glutamate transporters (3, 4). Recent studies have supported the glia as the major pathway of glutamate clearance (3). Neurons lack the enzymes necessary to perform net glutamate synthesis and depend on the glia to supply precursors. One of the pathways proposed for neuronal glutamate repletion is the glutamate͞glutamine cycle (5-8). In this pathway, glutamate taken up by the glia is converted to glutamine by glutamine synthetase (9-11). Glutamine then is released to the extracellular fluid, where it is taken up by neurons and is converted back to glutamate by the action of phosphate-activated glutaminase (12).The rate of the glutamate͞glutamine cycle has been controversial because of difficulties in performing measurements in the living brain. The prevailing belief has been that the glutamate͞glutamine cycle is a minor metabolic flux relative to total cellular glutamate metabolism. This view is largely based on the small size of the vesicular glutamate pool compared with other cellular glutamate pools (13,14). Additional evidence comes from the low flux of isotope from [1-13 C] glucose into glutamine in studies of brain slices (15).We have demonstrated that in vivo 13 C NMR may be used to measure the rate of glutamine labeling (16, 17) from [1-13 C] glucose in human occipital͞parietal cortex. These and subsequent studies (18) demonstrated that, in contrast with results from nonactivated brain slices (15), glutamine labeling is rapid. However, the rate of the glutamate͞glutamine cycle was not uniquely determined from these first experiments because of the inability to distinguish the glutamate͞glutamine cycle from other sources of glutamine labeling. The major alternate pathway of brain glutamine metaboli...